+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-24225 | ||||||||||||
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タイトル | Low resolution, multi-part 3D structure of the isolated yeast NPC | ||||||||||||
マップデータ | full NPC 3D map at 24A resolution | ||||||||||||
試料 |
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機能・相同性 | 機能・相同性情報 response to spindle checkpoint signaling / nuclear pore linkers / : / regulation of protein desumoylation / peroxisomal importomer complex / mRNA export from nucleus in response to heat stress / Seh1-associated complex / positive regulation of ER to Golgi vesicle-mediated transport / protein exit from endoplasmic reticulum / protein localization to nuclear inner membrane ...response to spindle checkpoint signaling / nuclear pore linkers / : / regulation of protein desumoylation / peroxisomal importomer complex / mRNA export from nucleus in response to heat stress / Seh1-associated complex / positive regulation of ER to Golgi vesicle-mediated transport / protein exit from endoplasmic reticulum / protein localization to nuclear inner membrane / COPII-mediated vesicle transport / COPII-coated vesicle budding / nuclear pore inner ring / nuclear pore localization / adenyl-nucleotide exchange factor activity / regulation of nucleocytoplasmic transport / regulation of TORC1 signaling / establishment of mitotic spindle localization / nuclear pore central transport channel / nuclear migration along microtubule / transcription-dependent tethering of RNA polymerase II gene DNA at nuclear periphery / telomere tethering at nuclear periphery / nuclear pore outer ring / nuclear pore complex assembly / nuclear pore organization / tRNA export from nucleus / COPII vesicle coat / nuclear pore cytoplasmic filaments / positive regulation of protein exit from endoplasmic reticulum / Transport of Mature mRNA derived from an Intron-Containing Transcript / post-transcriptional tethering of RNA polymerase II gene DNA at nuclear periphery / Regulation of HSF1-mediated heat shock response / nuclear pore nuclear basket / SUMOylation of SUMOylation proteins / cytoplasmic dynein complex / protein localization to kinetochore / SUMOylation of RNA binding proteins / structural constituent of nuclear pore / RNA export from nucleus / SUMOylation of chromatin organization proteins / nucleocytoplasmic transport / silent mating-type cassette heterochromatin formation / vacuolar membrane / regulation of mitotic nuclear division / poly(A)+ mRNA export from nucleus / nuclear localization sequence binding / dynein intermediate chain binding / NLS-bearing protein import into nucleus / cytoplasmic microtubule / establishment of mitotic spindle orientation / 加水分解酵素; プロテアーゼ; ペプチド結合加水分解酵素; セリンエンドペプチターゼ / ribosomal large subunit export from nucleus / positive regulation of TOR signaling / subtelomeric heterochromatin formation / mRNA transport / ribosomal small subunit export from nucleus / mRNA export from nucleus / heterochromatin formation / nuclear pore / ERAD pathway / positive regulation of TORC1 signaling / Neutrophil degranulation / cellular response to amino acid starvation / protein export from nucleus / nuclear periphery / molecular condensate scaffold activity / chromosome segregation / cell periphery / promoter-specific chromatin binding / phospholipid binding / protein import into nucleus / transcription corepressor activity / double-strand break repair / protein transport / nuclear envelope / single-stranded DNA binding / nuclear membrane / amyloid fibril formation / chromosome, telomeric region / hydrolase activity / cell cycle / cell division / chromatin binding / protein-containing complex binding / endoplasmic reticulum membrane / structural molecule activity / positive regulation of DNA-templated transcription / negative regulation of transcription by RNA polymerase II / endoplasmic reticulum / positive regulation of transcription by RNA polymerase II / DNA binding / RNA binding / identical protein binding / nucleus / cytoplasm / cytosol 類似検索 - 分子機能 | ||||||||||||
生物種 | Saccharomyces cerevisiae (パン酵母) | ||||||||||||
手法 | 単粒子再構成法 / クライオ電子顕微鏡法 / 解像度: 24.0 Å | ||||||||||||
データ登録者 | Akey CW / Rout MP / Ouch C / Echeverria I / Fernandez-Martinez J / Nudelman I | ||||||||||||
資金援助 | 米国, 3件
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引用 | ジャーナル: Cell / 年: 2022 タイトル: Comprehensive structure and functional adaptations of the yeast nuclear pore complex. 著者: Christopher W Akey / Digvijay Singh / Christna Ouch / Ignacia Echeverria / Ilona Nudelman / Joseph M Varberg / Zulin Yu / Fei Fang / Yi Shi / Junjie Wang / Daniel Salzberg / Kangkang Song / ...著者: Christopher W Akey / Digvijay Singh / Christna Ouch / Ignacia Echeverria / Ilona Nudelman / Joseph M Varberg / Zulin Yu / Fei Fang / Yi Shi / Junjie Wang / Daniel Salzberg / Kangkang Song / Chen Xu / James C Gumbart / Sergey Suslov / Jay Unruh / Sue L Jaspersen / Brian T Chait / Andrej Sali / Javier Fernandez-Martinez / Steven J Ludtke / Elizabeth Villa / Michael P Rout / 要旨: Nuclear pore complexes (NPCs) mediate the nucleocytoplasmic transport of macromolecules. Here we provide a structure of the isolated yeast NPC in which the inner ring is resolved by cryo-EM at sub- ...Nuclear pore complexes (NPCs) mediate the nucleocytoplasmic transport of macromolecules. Here we provide a structure of the isolated yeast NPC in which the inner ring is resolved by cryo-EM at sub-nanometer resolution to show how flexible connectors tie together different structural and functional layers. These connectors may be targets for phosphorylation and regulated disassembly in cells with an open mitosis. Moreover, some nucleoporin pairs and transport factors have similar interaction motifs, which suggests an evolutionary and mechanistic link between assembly and transport. We provide evidence for three major NPC variants that may foreshadow functional specializations at the nuclear periphery. Cryo-electron tomography extended these studies, providing a model of the in situ NPC with a radially expanded inner ring. Our comprehensive model reveals features of the nuclear basket and central transporter, suggests a role for the lumenal Pom152 ring in restricting dilation, and highlights structural plasticity that may be required for transport. | ||||||||||||
履歴 |
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-構造の表示
ムービー |
ムービービューア |
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構造ビューア | EMマップ: SurfViewMolmilJmol/JSmol |
添付画像 |
-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_24225.map.gz | 61.7 MB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-24225-v30.xml emd-24225.xml | 31.1 KB 31.1 KB | 表示 表示 | EMDBヘッダ |
FSC (解像度算出) | emd_24225_fsc.xml | 14.3 KB | 表示 | FSCデータファイル |
画像 | emd_24225.png | 211.9 KB | ||
その他 | emd_24225_additional_1.map.gz emd_24225_additional_2.map.gz emd_24225_additional_3.map.gz emd_24225_additional_4.map.gz emd_24225_half_map_1.map.gz emd_24225_half_map_2.map.gz | 66.4 MB 2.9 MB 66.1 MB 66.6 MB 84.8 MB 85 MB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-24225 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-24225 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_24225_validation.pdf.gz | 438.4 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_24225_full_validation.pdf.gz | 437.9 KB | 表示 | |
XML形式データ | emd_24225_validation.xml.gz | 18.3 KB | 表示 | |
CIF形式データ | emd_24225_validation.cif.gz | 23.8 KB | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-24225 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-24225 | HTTPS FTP |
-関連構造データ
-リンク
EMDBのページ | EMDB (EBI/PDBe) / EMDataResource |
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「今月の分子」の関連する項目 |
-マップ
ファイル | ダウンロード / ファイル: emd_24225.map.gz / 形式: CCP4 / 大きさ: 113.6 MB / タイプ: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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注釈 | full NPC 3D map at 24A resolution | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
ボクセルのサイズ | X=Y=Z: 5.312 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
詳細 | EMDB XML:
CCP4マップ ヘッダ情報:
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-添付データ
-追加マップ: nucleoplasmic outer ring-focused C8 refinement
ファイル | emd_24225_additional_1.map | ||||||||||||
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注釈 | nucleoplasmic outer ring-focused C8 refinement | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-追加マップ: masked 3D map of central transporter
ファイル | emd_24225_additional_2.map | ||||||||||||
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注釈 | masked 3D map of central transporter | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-追加マップ: inner spoke ring- focused C8 refinement
ファイル | emd_24225_additional_3.map | ||||||||||||
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注釈 | inner spoke ring- focused C8 refinement | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-追加マップ: flexible cytoplasmic outer ring-focused C8 refinement
ファイル | emd_24225_additional_4.map | ||||||||||||
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注釈 | flexible cytoplasmic outer ring-focused C8 refinement | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: half map full NPC
ファイル | emd_24225_half_map_1.map | ||||||||||||
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注釈 | half map full NPC | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-ハーフマップ: half map full NPC
ファイル | emd_24225_half_map_2.map | ||||||||||||
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注釈 | half map full NPC | ||||||||||||
投影像・断面図 |
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密度ヒストグラム |
-試料の構成要素
-全体 : 3D map of yeast NPC
全体 | 名称: 3D map of yeast NPC |
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要素 |
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-超分子 #1: 3D map of yeast NPC
超分子 | 名称: 3D map of yeast NPC / タイプ: complex / ID: 1 / 親要素: 0 詳細: low resolution 3D map from 1-step affinity isolation of NPCs, multibody and focused refinements |
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分子量 | 実験値: 52.0 MDa |
-超分子 #2: inner spoke ring of yeast NPC
超分子 | 名称: inner spoke ring of yeast NPC / タイプ: complex / ID: 2 / 親要素: 1 / 詳細: focused 3D map of inner ring |
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由来(天然) | 生物種: Saccharomyces cerevisiae (パン酵母) / Organelle: nucleus / 細胞中の位置: nuclear envelope |
-超分子 #3: double nuclear outer ring
超分子 | 名称: double nuclear outer ring / タイプ: complex / ID: 3 / 親要素: 1 / 詳細: 3D map visualized by focused C8 refinement |
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由来(天然) | 生物種: Saccharomyces cerevisiae (パン酵母) / Organelle: nucleus / 細胞中の位置: nuclear envelope |
-超分子 #4: central transporter
超分子 | 名称: central transporter / タイプ: complex / ID: 4 / 親要素: 1 / 詳細: masked 3D map of central channel density in NPC |
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由来(天然) | 生物種: Saccharomyces cerevisiae (パン酵母) / Organelle: nucleus / 細胞中の位置: nuclear envelope |
-超分子 #5: Nup82 complex hook ring
超分子 | 名称: Nup82 complex hook ring / タイプ: complex / ID: 5 / 親要素: 1 / 詳細: focused 3D density map of cytoplasmic outer ring |
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由来(天然) | 生物種: Saccharomyces cerevisiae (パン酵母) / Organelle: nucleus / 細胞中の位置: nuclear envelope |
-実験情報
-構造解析
手法 | クライオ電子顕微鏡法 |
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解析 | 単粒子再構成法 |
試料の集合状態 | particle |
-試料調製
濃度 | 0.3 mg/mL | |||||||||||||||||||||
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緩衝液 | pH: 7.5 構成要素:
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グリッド | モデル: Quantifoil R2/2 / 材質: COPPER / メッシュ: 400 / 支持フィルム - 材質: CARBON / 支持フィルム - トポロジー: CONTINUOUS / 支持フィルム - Film thickness: 5.0 nm / 前処理 - タイプ: GLOW DISCHARGE / 前処理 - 雰囲気: OTHER | |||||||||||||||||||||
凍結 | 凍結剤: ETHANE / チャンバー内湿度: 100 % / チャンバー内温度: 283 K / 装置: FEI VITROBOT MARK III 詳細: Grids were floated with carbon side down on 5 uL sample drops in a humid chamber. This was followed by 3 stepwise transfers onto 20 uL drops of blotting buffer while keeping the backside of ...詳細: Grids were floated with carbon side down on 5 uL sample drops in a humid chamber. This was followed by 3 stepwise transfers onto 20 uL drops of blotting buffer while keeping the backside of the grids dry. blot buffer was removed with a manual blot from the bottom edge of the grid through a side port in the humid chamber; an appropriate volume of blot buffer was pipetted immediately onto the grid, which was double blotted and plunge frozen.. | |||||||||||||||||||||
詳細 | NPCs were released gently from the NE with detergent-extraction of a cell cryo-lysate and purified with a single affinity step; to minimize local domain movements NPCs were mildly cross-linked by the addition of DiSuccinimidylSuberate |
-電子顕微鏡法
顕微鏡 | FEI TITAN KRIOS |
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特殊光学系 | エネルギーフィルター - 名称: GIF Bioquantum / エネルギーフィルター - スリット幅: 20 eV |
撮影 | フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 検出モード: SUPER-RESOLUTION / デジタル化 - サイズ - 横: 3838 pixel / デジタル化 - サイズ - 縦: 3710 pixel / デジタル化 - サンプリング間隔: 5.0 µm / デジタル化 - 画像ごとのフレーム数: 2-40 / 撮影したグリッド数: 1 / 実像数: 4015 / 平均露光時間: 0.5 sec. / 平均電子線量: 40.0 e/Å2 詳細: Navigator parameters were set to have 6-7 groups of holes per mesh, with one focus spot per group using an in-house script from Dr. Chen Xu. |
電子線 | 加速電圧: 300 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | C2レンズ絞り径: 70.0 µm 最大 デフォーカス(補正後): 3.8000000000000003 µm 最小 デフォーカス(補正後): 1.5 µm / 倍率(補正後): 37651 / 照射モード: OTHER / 撮影モード: BRIGHT FIELD / Cs: 2.7 mm / 倍率(公称値): 50000 |
試料ステージ | ホルダー冷却材: NITROGEN |
実験機器 | モデル: Titan Krios / 画像提供: FEI Company |